State Key Laboratory of Southwestern Chinese Medicine Resources, Ministry of Education, Chengdu 611137, China.
School of Pharmacy, Chengdu University of Traditional Chinese Medicine, Chengdu 611137, China.
Oxid Med Cell Longev. 2022 Jul 16;2022:4591134. doi: 10.1155/2022/4591134. eCollection 2022.
Alcoholic liver disease (ALD) is a multifaceted process that involves excessive lipid, reactive oxygen species (ROS) production, unbalanced mitochondrial homeostasis, and ultimate cell death. Quercetin is a dietary phytochemical presented in various fruits and vegetables, which has anti-inflammatory and antioxidant effects. According to recent advances in pharmanutritional management, the effects of quercetin on various mitochondrial processes have attracted attention. In the study, we explored whether quercetin could attenuate ethanol-induced hepatocyte pyroptosis by maintaining mitochondrial homeostasis and studied its hepatoprotective effect and the underlying mechanism. We chose L02 cells to establish an model with ethanol-induced hepatocyte pyroptosis. Then, the cells at approximately 80% confluence were treated with quercetin (80, 40, and 20 M). The cell viability (CCK-8) was used to investigate the effect of quercetin on ethanol-induced L02 cell proliferation. Relative assay kits were used to measure oxidative stress index (OSI = TOS/TAS), lipid peroxidation (LPO) release, and mitochondrial membrane potential (Ψ). The morphology of mitochondria was characterized by transmission electron microscopy- (TEM-) based analysis. Mitochondrial dynamics (Mito Tracker Green), mitROS (MitoSOX Red Mitochondrial Superoxide) production, and nuclear DNA (nDNA) damage (H2AX) markers were detected by immunofluorescence. The mRNA levels of mitochondrial function (including mitochondrial DNA (mtDNA) transcription genes TWNK, MTCO1, and MFND) and pyroptosis-related genes were detected by RT-qPCR, and the protein levels of NLRP3, ASC, caspase1, cleaved-caspase1, IL-18, IL-1, and GSDMD-N were detected by western blot. The results showed that quercetin treatment downregulated redox status, lipid droplets, and LPO release, restored damaged mitochondrial membrane potential, and repaired mtDNA damage, PGC-1 nuclear transfer, and mitochondrial dynamics. The gene and protein expressions of NLRP3, ASC, cleaved-caspase1, IL-18, IL-1, and GSDMD-N were decreased, which effectively inhibited cell pyroptosis. Therefore, the results indicated that quercetin protected ethanol-induced hepatocyte pyroptosis via scavenging mitROS and promoting PGC-1-mediated mitochondrial homeostasis in L02 cells.
酒精性肝病（ALD）是一个多方面的过程，涉及脂质、活性氧（ROS）的过度产生、线粒体平衡的破坏以及最终的细胞死亡。槲皮素是一种存在于各种水果和蔬菜中的膳食植物化学物质，具有抗炎和抗氧化作用。根据 Pharmanutritional 管理的最新进展，槲皮素对各种线粒体过程的影响引起了人们的关注。在这项研究中，我们探讨了槲皮素是否可以通过维持线粒体平衡来减轻乙醇诱导的肝细胞细胞焦亡，并研究了其肝保护作用及其潜在机制。我们选择 L02 细胞建立乙醇诱导的肝细胞细胞焦亡模型。然后，将大约 80%汇合的细胞用槲皮素（80、40 和 20μM）处理。用 CCK-8 法检测槲皮素对乙醇诱导的 L02 细胞增殖的影响。相对测定试剂盒用于测量氧化应激指数（OSI = TOS/TAS）、脂质过氧化（LPO）释放和线粒体膜电位（Ψ）。用透射电子显微镜（TEM）分析来描述线粒体的形态。通过免疫荧光法检测线粒体动力学（MitoTracker Green）、mitROS（MitoSOX Red Mitochondrial Superoxide）的产生和核 DNA（nDNA）损伤（H2AX）标志物。用 RT-qPCR 检测线粒体功能（包括线粒体 DNA（mtDNA）转录基因 TWNK、MTCO1 和 MFND）和细胞焦亡相关基因的 mRNA 水平，用 Western blot 检测 NLRP3、ASC、caspase1、cleaved-caspase1、IL-18、IL-1 和 GSDMD-N 的蛋白水平。结果表明，槲皮素处理下调了氧化还原状态、脂滴和 LPO 释放，恢复了受损的线粒体膜电位，修复了 mtDNA 损伤、PGC-1 核转移和线粒体动力学。NLRP3、ASC、cleaved-caspase1、IL-18、IL-1 和 GSDMD-N 的基因和蛋白表达降低，有效抑制了细胞焦亡。因此，结果表明，槲皮素通过清除 mitROS 和促进 PGC-1 介导的 L02 细胞线粒体平衡来保护乙醇诱导的肝细胞细胞焦亡。
